[Chemical reaction mechanism of decoction of traditional Chinese medicines: a review].

Complicated chemical reactions occur in the decoction of traditional Chinese medicines(TCMs) which features complex components, influencing the safety, efficacy, and quality controllability of TCMs. Therefore, it is particularly important to clarify the chemical reaction mechanism of TCMs in the decoction. This study summarized eight typical chemical reactions in the decoction of TCMs, such as substitution reaction, redox reaction, isomerization/stereoselective reaction, complexation, and supramolecular reaction. With the "toxicity attenuation and efficiency enhancement" of aconitines and other examples, this study reviewed the reactions in decoction of TCMs, which was expected to clarify the variation mechanisms of key chemical components in this process and to help guide medicine preparation and safe and rational use of medicine in clinical settings. The current main research methods for chemical reaction mechanisms of decoction of TCMs were also summed up and compared. The novel real-time analysis device of decoction system for TCMs was found to be efficient and simple without the pre-treatment of samples. This device provides a promising solution, which has great potential in quantity evaluation and control of TCMs. Moreover, it is expected to become a foundational and exemplary research tool, which can advance the research in this field.

Research progress of quality control for the seed of Ziziphus jujuba var. spinosa (Bunge) Hu ex H.F. Chow (Suan-Zao-Ren) and its proprietary Chinese medicines.

ETHNOPHARMACOLOGICAL RELEVANCE: Semen Ziziphi Spinosae (SZS), the seed of Ziziphus jujuba var. spinosa (Bunge) Hu ex H.F. Chow (Chinese name Suan-Zao-Ren), is widely distributed in China, Laos, Myanmar, and Iran. It is a classic traditional Chinese medicine with sedative and sleeping effects. In clinical practice, there are more than 155 proprietary Chinese medicines containing SZS. However, many commercial SZS products are difficult to qualify using current methods. Moreover, there is a scarcity of quality standards for SZS in proprietary Chinese medicines. AIM OF THE STUDY: The purpose of this study was to clearly reveal the quality indicators during the entire production process of SZS and its products. MATERIALS AND METHODS: This study reviewed more than 230 articles and related books on the quality control of SZS and its proprietary Chinese medicines published over the last 40 years (from January 1979 to October 2022). Moreover, where available, information on the quality of SZS and its proprietary Chinese medicines was also collected from websites for comparison, including online publications (e.g. PubMed, CNKI, Google Scholar, and Web of Science), the information at Yaozhi website and China Medical Information Platform, along with some classic books on Chinese herbal medicine. The literature and information search were conducted using keywords such as "Suan-Zao-Ren", " Ziziphus jujuba" and "quality control", and the latest results from various databases were combined to obtain valid information. The active components, which in vivo exposure, and Q-markers were also summarized. RESULTS: The jujuboside A, jujuboside B, and spinosin were revealed as the key Q-markers for SZS. Moreover, the advancements and prospects of the quality control for SZS and its extract, proprietary Chinese medicines, health foods, and adulterants were comprehensively summarized. The high-performance liquid chromatography-UV/evaporative light scattering detection and fingerprint analysis were found to be the mainstream methods for the SZS quality control. In particular, the novel quality evaluation method based on the unit content was applied for SZS and its proprietary Chinese medicines. Significant fluctuations were found in the contents of Q-markers. Moreover, the mass transfer rule of Q-markers was comprehensively clarified based on the entire production process, including production origins, ripening time, primary process, processing, compatibility decoction/extract, and storage. Ultimately, the crushing and compatibility of SZS were found to be the key steps affecting the active components. CONCLUSIONS: In short, this study provides solid evidences to reveal quality indicators for the entire production process of developing rational quality standards for SZS and its products. Moreover, this study also provides a template quality control overview, which could be extended to other traditional Chinese medicines.

Compatibility of Fuzi and Ginseng Significantly Increase the Exposure of Aconitines.

The herb-pair ginseng-Fuzi (the root of Aconitum carmichaelii) is the material basis of Shenfu prescriptions and is popular in traditional Chinese medicine for the treatment of heart failure, and even shock with severe-stage of COVID-19. A narrow therapeutic window of Fuzi may cause significant regional loss of property and life in clinics. Therefore, systemic elucidation of active components is crucial to improve the safety dose window of Shenfu oral prescriptions. A high performance liquid chromatography-mass spectrometry method was developed for quantification of 10 aconitines in SD rat plasma within 9 min. The limit of detection and the limit of quantification were below 0.032 ng/ml and 0.095 ng/ml, respectively. Furthermore, a systemic comparison with their pharmacokinetic characteristics after oral administration of a safe dosage of 2 g/kg of Fuzi and ginseng-Fuzi decoction for 24 h was conducted. Eight representative diester, monoester, and non-ester aconitines and two new active components (i.e., songorine and indaconitine) were all adopted to elucidating the differences of the pharmacokinetic parameters in vivo. The compatibility of Fuzi and ginseng could significantly increase the in vivo exposure of active components. The terminal elimination half-life and the area under the concentration-time curve of mesaconitine, benzoylaconitine, benzoylmesaconitine, benzoylhypaconitine, and songorine were all increased significantly. The hypaconitine, benzoylmesaconitine, and songorine were regarded as the main active components in vivo, which gave an effective clue for the development of new Shenfu oral prescriptions.

Discovery of the directionally detoxification effect and chemical mechanism of Ginseng-Fuzi co-decoction based on real-time online filtration electrospray ionization mass spectrometry.

BACKGROUND: The synergic action of compound prescriptions is an important feature and core advantage of traditional medicine. Ginseng-Fuzi decoction is a classic compatible phytomedicine in China, of which Ginseng can effectively reduce the toxicity of Fuzi in clinical, but the detoxification chemical mechanism is still unclear. PURPOSE: Develop a novel method for real-time tracking and monitoring of complex substances in the decoction system of traditional Chinese medicine to uncover the detoxification effect Ginseng on Fuzi and explore the possible chemical reaction mechanism of Ginseng-Fuzi co-decoction. METHODS: A novel real-time monitoring system, online filtration electrospray ionization mass spectrometry, was developed for extremely complex substances analysis in the decoction of traditional medicine compounds to uncover the directionally detoxification effect and the mechanism of compatibility interaction. RESULTS: Nine key alkaloids and 7 ginsenosides in Ginseng-Fuzi decoction were simultaneously in-situ monitoring in positive ion mode or negative ion mode respectively. Both types of targeted analytes had satisfactory MS signal response for real-time qualitative and quantitative analysis with high precision (RSD < 14.04%) and low LLODs (0.002 ng/ml-10 ng/ml). Through long-term tracking analysis, the exact detoxification and synergistic effect of Ginseng-Fuzi decoction were confirmed as the concentration of main toxic alkaloids decreased (e.g. the content of mesaconitine has been reduced by about 38%) and the main active monoester alkaloids increased obviously. More importantly, the possible molecular mechanism of the detoxification effect of Ginseng compatibility was revealed for the first time, which was the nucleophilic substitution reaction of diester alkaloids catalyzed by fatty acids. CONCLUSION: This study revealed the exact effect of co-decoction of Ginseng and Fuzi at the molecular level and the chemical reaction mechanism of fatty acid-catalyzed degradation of toxic diester-type alkaloids. The comprehensive multi-component real-time monitoring strategy for complex traditional medicine compounds developed and implemented here has important demonstration significance for revealing the scientific connotation of the compatibility of compound traditional medicine.

[Construction of quality evaluation strategy for Chinese medicine based on ambient mass spectrometry].

Quality is the guarantee for the clinical safety and effectiveness of Chinese medicine. Accurate quality evaluation is the key to the standardization and modernization of Chinese medicine. Efforts have been made in improving Chinese medicine quality and strengthening the quality and safety supervision in China, but rapid and accurate quality evaluation of complex Chinese medicine samples is still a challenge. On the basis of the development of ambient mass spectrometry and the application in quality evaluation of complex Chinese medicine systems in recent years, the authors developed the multi-scenario Chinese medicine quality evaluation strategies. A systematic methodology was proposed in specific areas such as real-time monitoring of the quality of complex Chinese medicine decoction system, rapid toxicity grading of compound Chinese patent medicine, and evaluation of bulk medicinals of Chinese patent medicine. Allowing multi-scenario analysis of Chinese medicine, it is expected to provide universal research ideas and technical methods for rapid and accurate quality evaluation of Chinese medicine and boost the high-quality development of Chinese medicine industry.

Molecular differentiation of Panax notoginseng grown under different conditions by internal extractive electrospray ionization mass spectrometry and multivariate analysis.

Panax notoginseng is a highly valuable and widely used herb in traditional Chinese medicine. The quality and efficacy of Panax notoginseng grown under different conditions can greatly vary due to the differences in chemical composition. The analysis of chemical composition in Panax notoginseng typically involves various experimental steps including extraction, chromatographic separation and characterization, which can be time- and labor-consuming. Therefore, the efficient quality assessment and control of Panax notoginseng requires the development of more rapid methods for the chemical characterization and classification of Panax notoginseng. In this study, a method based on internal extractive electrospray ionization mass spectrometry (iEESI-MS) was developed to characterize chemical components of Panax notoginseng samples under different growth conditions (e.g., place of origin, soil quality, growth season) at the speed of 0.5 min per sample, without sample pretreatment and chromatographic separation. A total of 35 chemical components, including sugars, saponins, organic acids, etc., were identified in Panax notoginseng samples. Clear separation was observed in the multivariate analysis of the iEESI-MS data from Panax notoginseng samples grown under different conditions. The difference in the content of sucrose, fructose, Rg1, Rf, Rb1, Noto-R1, malonyl-Rb1, malonyl-Rg1, malonyl-Rf, Rd, Re, linoleic acid, palmitic acid and malic acid can be used as key characteristic indicators to discriminate origin, commercial specifications, and cultivation conditions of Panax notoginseng samples. The results of our study indicate the high power of iEESI-MS for the rapid molecular characterization and classification of Panax notoginseng under different growth conditions, which can be used for the quality assessment of traditional herbal medicines as well as in pharmaceutical and clinical analysis.

Chemical-activity-based quality marker screening strategy for Viscum articulatum.

Viscum articulatum Burm. f. is a parasitic plant rich in flavonoids, triterpenoids, and catechins and has a high nutritional value. It has been reported that consuming V. articulatum can prevent cardiac diseases. In this study, six bioactive compounds, including catechins, triterpenoids, and phenylpropanoid glycosides, were determined in alcohol extracts of the plant using HPLC. The anti-inflammatory and antioxidant activities of three catechins, two triterpenoids, and three combination drugs were measured in cardiomyocytes, and the results showed that the anti-inflammatory activity was significantly enhanced while retaining strong antioxidant activity when epicatechin and ursolic acid were used in combination. The main quality markers epicatechin and ursolic acid were screened based on the specificity of the genuine herb and a potent synergistic effect, and the lowest limitation contents of V. articulatum which could discriminate it from some other taxonomically similar materials were accordingly determined. This self-built lowest limitation content of the two screened quality markers could quickly and accurately reflect the efficacy in terms of chemical composition and reverse the disorderly market use of nongenuine herbs or confusing species for adulteration. This study is of some significance for market regulation, drug development, and clinical medication.

Online discovery of the molecular mechanism for directionally detoxification of Fuzi using real-time extractive electrospray ionization mass spectrometry.

ETHNOPHARMACOLOGICAL RELEVANCE: Aconitum carmichaelii Debeaux, a famous traditional medicinal herb for collapse, rheumatic fever, and painful joints, always raises global concerns about its fatal toxicity from toxic alkaloids when improperly processed. Therefore, it is urgent to clarify the internal molecular mechanism of processing detoxification on Aconitum and develop simple and reliable approaches for clinical application, which is also of great significance to the rational medicinal use of Aconitum. AIM OF THE STUDY: The study aimed at developing a complete molecular mechanism exploration strategy in complex medicinal herb decocting system, clarifying the internal molecular mechanism of processing detoxification on Aconitum, and exploring valid approaches for detoxification. MATERIALS AND METHODS: Aconiti Lateralis Radix Praeparata (Fuzi) was selected as the model for exploring the complex Aconitum detoxification mechanism using an advanced online real-time platform based on extractive electrospray ionization mass spectrometry. The methods realized the sensitive capture of dynamic trace intermediates, accurate qualitative and quantitative analysis, and real-time and long-term monitoring of multi-components with satisfactory accuracy and resistance to complex matrices. RESULTS: Components in the complex Aconitum decocting system were real-timely characterized and fat meat was discovered and verified to directionally detoxify Aconitum while reserving the therapy effect. More importantly, the dynamic detoxification mechanism in the chemically complex Aconitum decoction was molecularly profiled. A novel reaction pathway based on nucleophilic substitution reaction mechanism was proposed. As confirmed by the theoretic calculations at DFT B3LYP/6-31G (d) levels, fatty acids (e.g., palmitic acid) acted as a green, cheap, and high-performance catalyst and promote the decomposition of toxic diester alkaloids to non-toxic and active benzoyl-monoester alkaloids through the discovered mechanism. CONCLUSION: The study exposed a novel detoxification molecular mechanism of Aconitum and provided an effective method for the safe use of Aconitum, which could effectively guide the development of traditional processing technology and compatibility regulation of the toxic herb and had great value to the modernization and standardization development of traditional medicine.

Exploring the resources of the genus Viscum for potential therapeutic applications.

ETHNOPHARMACOLOGICAL RELEVANCE: The genus Viscum comprises approximately 100 species that are mainly distributed across Africa, Asia and Europe. The extracts and preparations of Viscum species are widely used as common complementary and alternative medicines in the treatment of rheumatism and cancer. AIM OF THE REVIEW: This review aims to explore the medicinal properties of twelve species belonging to the genus Viscum for potential therapeutic applications. MATERIALS AND METHODS: We collected online information (including PubMed, CNKI, Google Scholar, and Web of Science) from January 1915 to April 2021 and knowledge from classical books on Chinese herbal medicines available for 12 species of the genus Viscum, including Viscum coloratum (Kom.) Nakai, Viscum album L., Viscum articulatum Burm. f., Viscum liquidambaricola Hayata, Viscum ovalifolium DC., Viscum capitellatum Sm., Viscum cruciatum Sieber ex Boiss., Viscum nudum Danser, Viscum angulatum B.Heyne ex DC., Viscum tuberculatum A.Rich., Viscum multinerve Hayata, and Viscum diospyrosicola Hayata. RESULTS: At least 250 different compounds have been reported across twelve Viscum species, including amino acid and peptides, alkaloids, phenolic acids, flavonoids, terpenoids, carbohydrates, fatty acids, lipids, and other types of compounds. In particular, for Viscum coloratum (Kom.) Nakai and Viscum album L., the plants, preparations, and bioactive components have been thoroughly reviewed. This has allowed to elucidate the role of active components, including lectins, viscotoxins, flavonoids, terpenoids, phenolic acids, and polysaccharides, in multiple bioactivities, such as anti-cancer, anti-rheumatism arthralgia, anti-inflammation, anti-cardiovascular diseases, enhancing immunity, and anti-chemotherapy side effects. We also evaluated quality control methods based on active compounds, in vivo exposure compounds, and discriminated chemical markers. CONCLUSIONS: This is the first report to systematically review the pharmaceutical development history, chemical composition, clinical evidence, pharmacological activity, discriminated chemical markers, in vivo exposure, and quality control on twelve distinct species of Viscum plants with medicinal properties. The significant safety and efficacy, along with the minor side effects are constantly confirmed in clinics. The genus Viscum is thus an important medicinal resource that is worth exploring and developing in future pharmacological and chemical studies.

[Research progress on identification methods of growth years of traditional Chinese medicinal materials].

The growth years of traditional Chinese medicinal materials are closely related to their quality, which directly affects the efficacy and safety of clinical medication. Therefore, it is particularly important to establish an identification method for the growth years of traditional Chinese medicinal materials. In this review, the identification methods for the growth years of traditional Chinese medicinal materials were summarized systematically, and were divided into four types according to the identification principles and methods: traditional identification, molecular identification, physical/chemical identification, and integrated identification. Relying on rich experience, objective molecular markers, various physical/chemical methods and integrated identification techniques(including infrared spectroscopy, nuclear magnetic resonance spectroscopy, high performance liquid chromatography, gas chromatography, mass spectrometry, bionic identification technology and their tandem technologies, etc.), the differences of characters or chemical fingerprints were compared in depth. The growth years of traditional Chinese medicinal materials were quickly identified or predicted by the appearance and characters, the whole fingerprint information or the content of specific chemical markers, and their content ratios. Through the case analysis of mature varieties, we intend to promote the establishment of a perfect technology system for the identification of the growth years of traditional Chinese medicinal materials, and to provide a reference for other perennial herbal materials, finally resulting in the accurate and precise quality control of traditional Chinese medicinal materials.

Quality tracing evaluation strategies of compatible materials in Aconitum proprietary Chinese medicines.

The proprietary Chinese medicine (PCM) has become a significant supplement of modern medicine. Nevertheless, the absence of quality control standard of compatible materials in PCM has led to serious adulteration, which has an extremely bad effect on safety of drug use and clinical efficacy. Here, a quality tracing evaluation strategy of compatible materials in 32 Aconitum proprietary Chinese medicines (APCMs) was established, including data normalization, model development, model verification, and unknown prescription cracking. The model was delimited based on the weighted content of total 9 key alkaloids in 24 APCMs, which were 5.65-57.10 µg/g for extract medicines and 42.62-380.61 µg/g for powder medicines. Three newly published commercial APCMs, including Wangbi Tablet, Wangbi Granule, and Fengshigutong Capsule, were used to verify its reliability and the results proved to be positive. Moreover, a novel prescription cracking approach was proposed to decode the content of each material in five unknown prescriptions including Yaoxitong Capsule, Tongrendahuoluo Pill, Xinbao Pill, Dahuoluo Capsule, and Mugua Pill. Ultimately, the single or two compatible Aconitum materials in APCMs was successfully decoded and the processed level of the materials were effectively judged. This study for the first time established a practical strategy for supervision and cracking of compatible materials in PCMs and is of great significance to improve the quality control of PCMs.

Limitation standard of toxic aconitines in Aconitum proprietary Chinese medicines using on-line extraction electrospray ionization mass spectrometry.

Development of rapid analytical methods and establishment of toxic component limitation standards are of great importance in quality control of traditional Chinese medicine. Herein, an on-line extraction electrospray ionization mass spectrometry (oEESI-MS) coupled with a novel whole process integral quantification strategy was developed and applied to direct determination of nine key aconitine-type alkaloids in 20 Aconitum proprietary Chinese medicines (APCMs). Multi-type dosage forms (e.g., tablets, capsules, pills, granules, and liquid preparation) of APCM could be determined directly with excellent versatility. The strategy has the characteristics of high throughput, good tolerance of matrix interference, small amount of sample (∼0.5 mg) and reagent (∼240 µL) consumption, and short analysis time for single sample (<15 min). The results were proved to be credible by high performance liquid chromatography-mass spectrometry (LC-MS) and electrospray ionization mass spectrometry, respectively. Moreover, the limitation standard for the toxic aconitines in 20 APCMs was established based on the holistic weight toxicity (HWT) evaluation and the Chinese Pharmacopoeia severally, and turned out that HWT-based toxicity evaluation results were closer to the real clinical applications. Hence, a more accurate and reliable APCM toxicity limitation was established and expected to play an important guiding role in clinics. The current study extended the power of ambient MS as a method for the direct quantification of molecules in complex samples, which is commonly required in pharmaceutical analysis, food safety control, public security, and many other disciplines.

Real-time toxicity prediction of Aconitum stewing system using extractive electrospray ionization mass spectrometry.

Due to numerous obstacles such as complex matrices, real-time monitoring of complex reaction systems (e.g., medicinal herb stewing system) has always been a challenge though great values for safe and rational use of drugs. Herein, facilitated by the potential ability on the tolerance of complex matrices of extractive electrospray ionization mass spectrometry, a device was established to realize continuous sampling and real-time quantitative analysis of herb stewing system for the first time. A complete analytical strategy, including data acquisition, data mining, and data evaluation was proposed and implemented with overcoming the usual difficulties in real-time mass spectrometry quantification. The complex Fuzi (the lateral root of Aconitum)-meat stewing systems were real-timely monitored in 150 min by qualitative and quantitative analysis of the nine key alkaloids accurately. The results showed that the strategy worked perfectly and the toxicity of the systems were evaluated and predicated accordingly. Stewing with trotters effectively accelerated the detoxification of Fuzi soup and reduced the overall toxicity to 68%, which was recommended to be used practically for treating rheumatic arthritis and enhancing immunity. The established strategy was versatile, simple, and accurate, which would have a wide application prospect in real-time analysis and evaluation of various complex reaction systems.

[Microecology and geoherbalism of traditional Chinese medicine].

Microecology was directly or indirectly involved in the growth and development, metabolism process, and component accumulation of traditional Chinese medicine(TCM) in various ways, which affected the formation and changes of the geoherbalism of TCM. It was one of the main tasks of traditional Chinese medical microecology(TCMM) to reveal the relationship among microecological structure and its change rule and the quality effect of TCM. The heterogeneity of soil environment caused by geographical and climatic factors, as well as the discreteness limitation caused by isolation factors such as distance and host selection, were the main causes of the differentiation of microecological geography of TCM. The microecology of TCM had important influences and contributions on the distinctive origin and quality of Dao-di herbs, which was mainly reflected in the formation of excellent germplasm(including disease and insect resistance, drought resistance, salt resistance, cold resistance, etc.), the increase of yield, the formation of medicinal parts, the metabolism and accumulation of effective components, the time limit of harvesting, and the toxicity, increasing efficiency or reducing toxicity of TCM in the processing, the changes of product efficiency after introduction, and the authenticity of fungus medicine. With the vigorous development of metabonomics and modern information technology, the following aspects would become the future research trends, including the microecologically mediated biogenic pathway of chemical components, the metabolic synthesis reactor of TCM based on the microecological quantitative effect relationship, the cultivation of genuine Chinese medicine based on reconstruction of microecological structure, the origin identification barcode traceability technology, and the toxicity reduction and efficiency enhancement technology of TCM based on the microecological.

[Design and application of quality standard of Dao-di herbs].

Dao-di herbs, which are widely recognized as medicinal materials with a high quality and good efficacy in clinic, are now facing the dilemma of absence of standard. This study focused on a pivotal scientific problem of design and application of quality standard of Dao-di herbs, and systematically illustrated the general rules for the quality standard of Dao-di herbs involving "four rules, six core contents, and three key methods". The quality standard of Dao-di herbs shall be fully based on literatures as well as habitat, planting/breeding, processing, characters, chemical-pharmacological/toxic data. The common requirements for the quality standard of Dao-di herbs contain "clear source, explicit origin, rational indicator, gradable quality, and multiple detection methods". Notably, traditional experiences and modern techniques, quality tracing management system, "quality determination by distinguishing characters" method, rapid detection technology, effective/toxic substances control method, were comprehensively applied in this standard to purse the objectification, automation, and intellectualization of detection technology. Appearance characters, chemical components, and bioactive parameters, unified effective/toxic indicators, quality markers, and pharmacopeial control indicators and reasonable ranges were included in rigorous quality standards for Dao-di herbs. Besides, simple grading method shall be developed to guide the implementation of "high quality-high price" policy. Eventually, the new quality standards for Dao-di herbs will lead international standards and promote the high-quality development of Dao-di herbs.

[Research advancement in mechanisms of processing and compatibility for detoxication of Aconitums].

Aconitums,represented by Aconite Radix,Aconiti Lateralis Radix Praeparata and Aconiti Kusnezoffh Folium,is a kind of traditional Chinese medicine with a long medicinal history in China. They possess the significant toxicity and therapeutic effects simultaneously. Their potent effects of rescuing from dying,curing rheumatism,anti-inflammation,and analgesia make Aconitums highly regarded by physicians and pharmacists of various dynasties. However,countless poisoning cases caused by an irrational use of Aconitums were reported. In case of improper application and exceeding the therapeutic window,the acute cardiotoxicity and neurotoxicity would be caused,seriously threatening health and even life of the users. Therefore,the clinical application of Aconitums is limited to some extent. To avoid its toxicity and ensure the safety of medicinal use,Aconitums is usually used in a form of its processed products instead of the crude herbs,or combined with some other traditional Chinese medicines in a normal prescription. A proper processing and compatibility method can detoxicate its severe toxicity,reduce the adverse reactions,and also significantly broaden the indications and application range of Aconitums. This provides a guarantee for the secondary exploitation and utilization of Aconitums. In this paper,the traditional processing methods of Aconitums,along with the modern advancement were reviewed,and the mechanisms of detoxification by processing and compatibility were also illuminated. The physical detoxification mode and chemical detoxification mode were found as two main detoxification ways for Aconitums. In particular,the detoxification by hydrolysis,ion-pair,and saponification were three main means. The mechanisms illustrated in this paper can be a reference to the development of modern processing method and a guidance for appropriate use of Aconitums in clinical application.

Characterization of polymeric phenolic acids and flavonoids in Clerodendranthi Spicati Herba using ultrahigh-performance liquid chromatography coupled to quadrupole time-of-flight tandem mass spectrometry with target and nontarget data mining strategy.

RATIONALE: Clerodendranthi Spicati Herba (CSH) is often used to treat urinary stones, urinary tract infections and nephritis in China. Much literature has reported that polymeric phenolic acids and flavonoids are the major bioactive components in CSH. Therefore, it is very meaningful to identify the polymeric phenolic acids and flavonoids in CSH. METHODS: Ultrahigh-performance liquid chromatography coupled to quadrupole time-of-flight tandem mass spectrometry (UHPLC/QTOF-MS/MS) analysis with target and nontarget data mining strategy was proposed to rapidly profile the polymeric phenolic acids and flavonoids in CSH. Diagnostic product ions and neutral loss filter were beneficial for identifying the polymeric phenolic acids and flavonoids from complex compounds in CSH. RESULTS: A total of 118 compounds, including 85 polymeric phenolic acids and 33 flavonoids, were reasonably identified in CSH by comparing their main fragmentation pathways with literature data, and 85 of them were discovered in CSH firstly by nontarget analysis. Nine potential compounds were characterized tentatively as new pentameric and hexameric phenolic acids in CSH. Six types of polymeric phenolic acids (monomer, dimer, trimer, tetramer, pentamer and hexamer) and four types of flavonoids (apigenin, kaempferol, luteolin and quercetin) were identified in CSH. CONCLUSIONS: The results indicated that the UHPLC/QTOF-MS/MS method coupled with target and nontarget data mining strategy was feasible and rational for identifying the polymeric phenolic acids and flavonoids in complex chemical constituents of CSH. The findings will be conducive to the discovery of the active ingredients of CSH and the establishment of quality standards.

Functional characterization of three flavonoid glycosyltransferases from Andrographis paniculata.

Andrographis paniculata is an important traditional medicinal herb in South and Southeast Asian countries with diverse pharmacological activities that contains various flavonoids and flavonoid glycosides. Glycosylation can transform aglycones into more stable, biologically active and structurally diverse glycosides. Here, we report three glycosyltransferases from the leaves of A. paniculata (ApUFGTs) that presented wide substrate spectra for flavonoid glycosylation and exhibited multi-site glycosylation on the substrate molecules. They acted on the 7-OH position of the A ring and were able to glycosylate several other different types of compounds. The biochemical properties and phylogenetic analysis of these glycosyltransferases were also investigated. This study provides a basis for further research on the cloning of genes involved in glycosylation from A. paniculata and offers opportunities for enhancing flavonoid glycoside production in heterologous hosts. These enzymes are expected to become effective tools for drug discovery and for the biosynthesis of derivatives via flavonoid glycosylation.

Elucidation of metabolite isomers of Leonurus japonicus and Leonurus cardiaca using discriminating metabolite isomerism strategy based on ultra-high performance liquid chromatography tandem quadrupole time-of-flight mass spectrometry.

The isomer structural discrimination is a significant challenge in metabolome analysis based on ultrahigh performance liquid chromatography tandem high-resolution mass spectrometry (UHPLC-HRMS). In this study, a new discriminating metabolite isomerism strategy is proposed to elucidate the metabolome, especially the isomers, of Leonurus japonicus and Leonurus cardiaca. This strategy consists of three steps. First, the metabolite biosynthesis pathways are constructed based on a home-built compound database to rapidly profile the compounds of interest using the multiple diagnostic product ions (DPIs) screening analysis and binary comparison based on SUMPRODUCT function. Second, the fragmentation patterns (e.g. the high-resolution DPIs, DPI ratios) and chromatographic elution order are defined based on scattered reference chromatographic and mass spectrometry data, calculated lipophilicity parameters, molecular hydrogen bond analysis, and chemical reference standards. Finally, all discovered isomerisms are mapped with the defined applicable rules and the isomers are identified conveniently. Using this strategy, a total of 257 compounds were tentatively characterized, including 212 potential novel compounds and 67 pairs of cis-, trans-, and positional isomers of flavonoids, phenylethanoid glycosides, glucaric acids, novel quinic acids, and esters of fatty acids. Moreover, 56 characteristic markers were identified to discriminate these two herbal medicines. This strategy may significantly improve the efficiency and reliability of identifying isomers found in metabolite biosynthesis pathways.

Comprehensive metabolic profile of phenolic acids and flavonoids in Glechomae Herba using ultra-high-performance liquid chromatography coupled to quadrupole-time-of-flight tandem mass spectrometry with diagnostic ion filtering strategy.

Glechomae Herba (GH), a traditional Chinese medicine, is commonly used for the treatment of urolithiasis and cholelithiasis based on its diuretic and cholagogic actions. However, relatively few studies have been conducted to describe this medicine's metabolites. Therefore, an ultra-high-performance liquid chromatography coupled to quadrupole time of flight tandem mass spectrometry (UHPLC-QTOF-MS/MS) analysis with diagnostic product ions and neutral loss filtering strategy was established for the metabolic profiling profile of GH. First, an analytical method was established to detect the compounds in GH extracts and complex rat biosamples, including urine, bile, plasma and feces, after oral administration of GH extracts based on the holistic fingerprint approach utilizing UHPLC-QTOF-MS/MS. Second, the exogenously sourced ions were filtered out by the MarkerView software incorporated in Microsoft Office. Third, the diagnostic product ions and neutral loss filtering strategy were developed for the rapid identification of the metabolites. Finally, a total of 91 constituents, including 13 prototype compounds and 78 metabolites, were unambiguously or reasonably identified in rat biosamples. The metabolites were mainly eliminated by urine (82 metabolites) and bile (19 metabolites), and notably few metabolites were eliminated through feces (4 metabolites). Glucuronidation, methylation and sulfation were the primary metabolic pathways of phenolic acids and flavonoids. The primarily urine and bile elimination of metabolites may be related to the anti-urolithiasis and anti-cholelithiasis effects of GH. This work demonstrated the feasibility of an integrated strategy for the rapid identification of the metabolites after oral administration of GH extracts provided essential data for exploring the bioactive ingredients of GH in the treatment of urolithiasis and cholelithiasis.